Synergistic Action of Silver and Zinc Oxide Nanoparticles Loaded with Ofloxacin for Targeted Treatment of Diphtheria

Diphtheria, caused by Corynebacterium diphtheriae, remains a global health concern due to periodic outbreaks and increasing antibiotic resistance. Conventional antibiotics like ofloxacin are often rendered less effective by bacterial defense mechanisms, including biofilm formation and efflux pumps. This study aims to investigate the synergistic antibacterial potential of silver (AgNPs) and zinc oxide (ZnO-NPs) nanoparticles co-loaded with ofloxacin against C. diphtheriae, and evaluate their efficacy, biofilm inhibition, and cytotoxicity. AgNPs and ZnO-NPs were synthesized using chemical reduction and sol-gel techniques, respectively, and loaded with ofloxacin. The formulations were characterized using TEM, FTIR, XRD, and zeta potential analysis. Antibacterial activity was assessed using zone of inhibition, MIC/MBC values, and biofilm inhibition assays. Cytotoxicity was evaluated using MTT assay on HEK293 cell lines. Statistical significance was determined using one-way ANOVA. The Ag-ZnO-Ofx formulation exhibited the highest antibacterial efficacy, with a 33 mm inhibition zone, MIC of 0.6 µg/mL, and 83% biofilm inhibition. The combination nanoformulation also demonstrated acceptable cytocompatibility (>82% viability). Statistical analysis revealed highly significant differences (p < 0.001) in antibacterial and anti-biofilm activities compared to free ofloxacin and monometallic formulations. Silver and zinc oxide nanoparticles co-loaded with ofloxacin significantly enhance antibacterial and antibiofilm effects against C. diphtheriae, providing a promising nanotherapeutic strategy for combating diphtheria and similar infections. Further in vivo studies are warranted to support clinical translation.